孕激素及脂寡糖在淋球菌致病中的机制研究

发布时间：2018-04-25 06:04

本文选题：淋球菌 + 孕激素　；参考：《华中科技大学》2016年博士论文

【摘要】：目的：研究孕激素对淋球菌诱导的NLRP3炎症蛋白复合体的调控方法：构建去卵巢化与正常BALB/c雌性小鼠淋球菌感染模型,将20只BALB/c小鼠随机分为4组：1.假手术+淋球菌接种组(intact);2.卵巢摘除+淋球菌接种组;3.卵巢摘除+孕激素(1mg/d)+淋球菌接种组；4.假手术+生理盐水接种组(control)。分别记录4组模型小鼠感染天数并检测接种后每日阴道排出的菌量、阴道内中性粒细胞数量,ELISA法检测接种后小鼠阴道内IL-1β、TNF-α及IL-6细胞因子表达水平,接种后第5日取小鼠阴道及宫颈组织,HE染色检测阴道及宫颈组织病理改变,免疫荧光法检测阴道组织NLRP3和caspase-1 p20水平。体外实验分为5组：1.THP-1细胞组(空白对照)；2.THP-1细胞+淋球菌组;3.THP-1细胞+淋球菌+10 ng/mL孕激素组；4.THP-1细胞+淋球菌+100 ng/mL孕激素组；5.THP-1细胞+淋球菌+1000 ng/mL孕激素组。ELISA法检测孕激素作用后淋球菌感染的THP-1细胞分泌IL-1β水平,western-blot法检测细胞内NLRP3 及 caspase-1水平,RT-PCR法检测细胞内IL-1β及NF-κB的mRNA水平,western-blot检测细胞核内NF-k B p65核转位,荧光探针法检测细胞内ROS含量。结果：1)体内实验：孕激素降低小鼠模型的淋球菌感染菌量,抑制感染淋球菌小鼠阴道及宫颈组织内炎症细胞浸润,降低IL-1β、TNF-α、IL-6等炎性因子及NLRP3、caspase-1 p20的表达：2)体外实验：孕激素抑制淋球菌诱导的THP-1细胞内NLRP3、caspase-1及NF-κB的活化,减少了细胞的IL-1β分泌及ROS的释放。结论：孕激素通过减少NF-κB的转录及ROS的产生抑制淋球菌诱导的NLRP3炎症体活化,从而减轻感染淋球菌小鼠生殖道中炎症细胞浸润,减少炎症因子的表达。该研究从另一个角度阐述女性淋病无症状感染机制,为淋病防治提供了新的策略。
[Abstract]:Objective: To study the regulation of gonococcal NLRP3 inflammatory protein complex induced by gonococci: to construct ovariectomized and normal BALB/c female mice with gonococcal infection, 20 BALB/c mice were randomly divided into 4 groups: 1. sham operation + gonococcal inoculation group (intact), 2. ovarian removal + gonococcal inoculation group, 3. ovariectomy + progesterone (1mg/d). + gonococcal inoculation group; 4. sham operation + physiological saline inoculation group (control). The number of infected days in 4 groups of mice was recorded and the amount of vaginal discharge, the number of neutrophils in vagina after inoculation, the expression level of IL-1 beta, TNF- A and IL-6 cell factor in vagina after inoculation were detected by ELISA method, and the vagina of mice was taken at fifth days after inoculation, Cervical tissue, HE staining and pathological changes of vagina and cervix, NLRP3 and caspase-1 P20 in vaginal tissue were detected by immunofluorescence. In vitro, 5 groups were divided into groups: 1.THP-1 cell group (blank control); 2.THP-1 cells + gonococcal group; 3.THP-1 cell + gonococcal +10 ng/mL gestation group; 4.THP-1 cells + gonococcal +100 ng/mL progestogen group 5.THP-1 cells + gonococcal +1000 ng/mL progesterone group.ELISA assay was used to detect the level of IL-1 beta in THP-1 cells infected by gonococcal infection. The level of NLRP3 and caspase-1 in cells was detected by Western-blot method. RT-PCR assay was used to detect the level of IL-1 beta and NF- kappa. The intracellular ROS content was detected by needle method. Results: 1) in vivo experiment: progestin decreased the amount of gonococcal infection in mice model, inhibited the infiltration of inflammatory cells in vaginal and cervical tissues of mice infected with gonococcus, reduced IL-1 beta, TNF- alpha, IL-6 and other inflammatory factors and the expression of NLRP3, caspase-1 P20: 2) in vitro experiment: progestin inhibition of gonococcus induced by gonococcus The activation of NLRP3, caspase-1 and NF- kappa B in THP-1 cells reduced the secretion of IL-1 beta and the release of ROS. Conclusion: progesterone reduces the activation of NLRP3 inflammatory body induced by gonococcus by reducing the transcription of NF- kappa B and ROS, thus reducing the infiltration of inflammatory cells in the genital tract of gonococcal mice and reducing the expression of inflammatory factors. From another point of view, the study expounds the mechanism of asymptomatic infection of gonorrhea in women, which provides a new strategy for prevention and treatment of gonorrhea.

【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R759.2

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